RNeasy Protect Animal Blood System

For collecting and stabilizing animal blood and purifying RNA and miRNA

Reliable stabilization of transcript levels at blood collection

Convenient collection and storage of small blood volumes

Rapid purification of high-quality total RNA

Effective recovery of miRNA

The RNeasy Protect Animal Blood System provides a complete solution for preparing high-quality RNA from animal blood. Blood from mouse, rat, and other small animals is conveniently collected in RNAprotect Animal Blood Tubes, which contain a reagent that immediately stabilizes cellular RNA. Total RNA is then purified using the RNeasy Protect Animal Blood Kit. Please note that the RNeasy Protect Animal Blood Kit and the RNAprotect Animal Blood Tubes are ordered separately.

Alternatively, the RNeasy Protect Animal Blood Kit can be used to obtain microRNA (miRNA), either in a total RNA fraction or in a separate small RNA fraction. For purification of total RNA containing miRNA, Buffer RWT (cat. no. 1067933) is also required. For purification of small RNA (containing miRNA) and large RNA in 2 separate fractions, the RNeasy MinElute Cleanup Kit (cat. no. 74204) is also required. The procedures can be fully automated on theQIAcube.

50 tubes for collection, storage, and transport of 500 µl animal blood samples with RNA stabilization; to be used with the RNeasy Protect Animal Blood Kit

The RNeasy Protect Animal Blood System is intended for molecular biology applications. This product is not intended for the diagnosis, prevention, or treatment of a disease.

Effective transcript stabilization.

Blood samples (500 μl) from the same rat were collected in RNAprotect Animal Blood Tubes or EDTA-containing tubes. Samples were stored at 15–25°C for 0 minutes (t=0) to 6 hours. RNA was purified using the RNeasy Protect Animal Blood Kit, and FOS expression was analyzed in duplicate by real-time RT-PCR. ΔCT on the Y-axis shows CT values subtracted from those for the t=0 samples. FOS expression remained stable in the RNAprotect stabilized samples, but changed with time in the EDTA-treated samples (equivalent results were observed with other rats; data not shown).

Reproducible RNA yields.

Blood was collected from 4 rats in RNAprotect Animal Blood Tubes (four 500 μl samples collected per rat), and RNA was purified using the RNeasy Protect Animal Blood Kit. RNA was quantified using a NanoDrop spectrophotometer. Variability in RNA yield (%CV) between blood samples from the same rat was between 2.4 and 5.9%; between rats, variability was 15%.

RNeasy Protect Animal Blood flowchart.

The RNeasy Protect Animal Blood System provides a complete solution for preparing high-quality RNA from animal blood. Blood from mouse, rat, and other small animals is conveniently collected in RNAprotect Animal Blood Tubes, which contain a reagent that immediately stabilizes cellular RNA. Total RNA is then purified using the RNeasy Protect Animal Blood Kit. Alternatively, the RNeasy Protect Animal Blood Kit can be used in combination with Buffer RWT to obtain total RNA, including miRNA.

Highly intact RNA.

Mouse blood (100 μl) was collected in RNAprotect Animal Blood Tubes and stored at 4°C for [A] 2 hours or [B] 2 weeks. RNA was purified using the RNeasy Protect Animal Blood Kit and analyzed on the Agilent 2100 Bioanalyzer. The RIN values were virtually the same, demonstrating preservation of RNA integrity.

Performance

Blood samples collected in RNAprotect Animal Blood Tubes can be conveniently shipped at ambient temperature or easily stored in a refrigerator. Samples can be stored at 15–25°C for up to 48 hours, at 2–8°C for up to 2 weeks, or at –20°C for at least 3 months with no significant RNA degradation (see figure "Highly intact RNA"). Collection of animal blood in RNAprotect Animal Blood Tubes prevents changes in transcript levels, ensuring reliable gene expression analysis in applications such as real-time RT-PCR and microarray analysis (see figure "Effective transcript stabilization"). The RNeasy Protect Animal Blood Kit delivers reproducible yields of RNA, and requires less than 30 minutes of hands-on time (see figure "Reproducible RNA yields"). Proven RNeasy technology delivers purified RNA that is highly pure, with virtually no genomic DNA contamination.

Principle

When blood samples are collected, the gene expression profile can change significantly within minutes. Anticoagulants such as EDTA prevent blood clotting, but do not prevent changes in gene expression due to induced transcription or regulated turnover of mRNA. Collection of animal blood in RNAprotect Animal Blood Tubes prevents changes in transcript levels. Following digestion and homogenization of the stabilized blood, RNA is then purified from the samples using well-established RNeasy silica-membrane technology (see figure "RNeasy Protect Animal Blood flowchart").

Procedure

Blood samples are collected in RNAprotect Animal Blood Tubes. The tubes contain a reagent that lyses blood cells and stabilizes intracellular RNA. The RNAprotect Animal Blood Tubes are centrifuged to pellet the samples, which are then washed with water and resuspended in Buffer RSB. After digestion in Buffer RBT with proteinase K, the samples are homogenized by centrifugation through QIAshredder spin columns. Ethanol is added to the samples, and the samples are then centrifuged through RNeasy MinElute spin columns. The bound total DNA is subjected to DNase digestion and washed with Buffer RW1, followed by Buffer RPE. Pure RNA is eluted in Buffer REB. The purification of RNA with the RNeasy Protect Animal Blood Kit can be fully automated on the QIAcube. A separate protocol is provided for isolation of total RNA including miRNA and requires wash Buffer RWT (ordered separately; cat. no. 1067933). For purification of small RNA (containing miRNA) and large RNA in 2 separate fractions, the RNeasy MinElute Cleanup Kit is also required.

Applications

Total RNA and miRNA purified with the RNeasy Protect Animal Blood System can be used for many downstream applications, including: